Coral-seeding devices with fish-exclusion features reduce mortality on the Great Barrier Reef

Restoration methods that seed juvenile corals show promise as scalable interventions to promote population persistence through anthropogenic warming. However, challenges including predation by fishes can threaten coral survival. Coral-seeding devices with refugia from fishes offer potential solutions to limit predation-driven mortality. In an 8-month field study, we assessed the efficacy of such devices for increasing the survival of captive-reared Acropora digitifera (spat and microfragments) over control devices (featureless and caged). Devices with fish-exclusion features demonstrated a twofold increase in coral survival, while most corals seeded without protection suffered mortality within 48 h. Overall, spat faced more grazing and higher mortality compared to microfragments, and upward-facing corals were more vulnerable than side-facing corals. Grazing-induced mortality varied by site, with lower activity in locations abundant in mat-forming cyanobacteria or Scleractinian corals. Many scraping parrotfish were found feeding on or near the seeded corals; however, bites by Scarus globiceps explained the most site-related variation in grazing. Cyanobacteria may be preferred over corals as a nutritional resource for scraping parrotfish—advancing our understanding of their foraging ecology. Incorporating side-facing refugia in seeding devices and deploying to sites with nutrient-rich food sources for fish are potential strategies to enhance coral survival in restoration programs.


Coral spawning, settlement, and grow out
Coral colonies of the species Acropora digitifera (Dana, 1846) were collected (permit G21/38,062.1 issued by the Great Barrier Reef Marine Park Authority [GBRMPA]) from Davies Reef (central, midshelf Great Barrier Reef [GBR]) prior to the 2021 Autumn (February to March) coral-spawning event.Following collection, colonies were transported to outdoor, flow-through seawater aquaria (average light intensity 74 μmol photons m −1 s −1 and temperature 27-28 °C) in the National Sea Simulator at the Australian Institute of Marine Science (AIMS, Townsville, Queensland) and maintained until spawning.The timing of spawning and the numbers of colonies that contributed to mass cultures are reported in Supplementary Table S1.Gamete bundles were collected, separated, washed, and fertilized as described in Pollock et al. 2017 51 .Embryos were then transferred to 500 L larval rearing tanks at a stocking density of ~ 0.5-1 larva mL −1 .Culture tanks received flowthrough 0.4 μm filtered seawater (FSW) at 27.5-28.0°C and gentle aeration began ~ 16 h post fertilization.Larvae remained in rearing tanks and once they reached settlement competency (~ 7-12 d, as determined by daily competency assays) they were settled en masse on coral rearing plugs.
Coral larvae were settled on conditioned aragonite plugs that hosted a mixed community of crustose coralline algae (CCA; e.g., Porolithon, Lithophyllum, Titanoderma; ; see Supplementary Material Sect.1.1) and biofilms to induce settlement.Aragonite coral "frag plugs" (20-mm Ø; Ocean Wonders) were used as coral larval settlement substrates.Clean frag plugs were placed in well-conditioned (mixed community of CCA and biofilms), semi-recirculating indoor aquaria (280 L, flow of 5 L min -1 , average Photosynthetic Active Radiation [PAR] of 160 μmol m -2 s -1 ) for 2-months prior to settlement.For settlement, 167 plugs were placed into clean, polyvinyl chloride (PVC) trays and trays were distributed to 50 L acrylic tanks (1 holding tray per tank, 12 trays, and 12 tanks in total).Approximately 3500 larvae (~ 20 larvae per plug) were added per tank and allowed to settle over 72 h.Settlement success on individual plugs was then assessed under a dissecting microscope and additional larvae (up to 1500) were added to tanks with low settlement.The maximum number of larvae added to tanks during settlement was 5000.Approximately 9 larvae settled per plug (range of 0 to 46 spat per plug), totaling ~ 18000 individual spat on 2000 plugs.
Settled spat were maintained in a semi-recirculating indoor aquarium (280 L, flow of 5 L min −1 ) under a daily feeding regime (unenriched Artemia [1 nauplius mL -1 ], mixed microalgae [2000 cells mL −1 ; Nannochloropsis oceania, Isochrysis sp., Chaetoceros muelleri, Dunaliella sp., Proteomonas sulcate], and enriched Rotifers [0.5 nauplii mL −1 ; Brachionus plicatilis, 60-180 micron; SELCO © S.parkle]) and a consistent light profile (PAR, initial 20 μmol m −2 s −1 and increasing ~ 14 μmol m −2 s −1 weekly, with a maximum of 160 μmol m −2 s −1 by deployment).PAR was maintained at a low level to control the growth of benthic organisms that compete with spat, and to mimic the low-light conditions found in crevices where spat tend to preferentially settle 29,36 .Fragments (> 10-cm branch length) of adult broodstock were placed in the aquarium with spat plugs to promote uptake of symbiotic zooxanthellae, Symbiodiniaceae.The aquarium was cleaned weekly, and plug trays were rotated and repositioned fortnightly to reduce the effect of any within-tank environmental variability on spat growth.After 4 months, 260 plugs were haphazardly distributed among 130 seeding devices for deployment, resulting in 2

Coral microfragmentation
Microfragments were generated from 5 adult broodstock colonies 3 months post spawning and 1 month prior to deployment.Colonies were first chiseled into large fragments (~ 10 cm length), then cut into uniform microfragments (8 × 8 mm, Gryphon Diamond Band Saw).Only branches with mature polyps (i.e., upward facing in the center of the colony) were chosen for microfragmentation 9 .Branches were first cross-sectioned; the upper facing side was then cut into microfragments.The microfragments were glued (Gorilla glue) to the center of clean (bleached and autoclaved) aragonite coral plugs (20 mm Ø) and maintained in aquaria for approximately 1 month for grow out.Microfragments (n = 692) on plugs were then distributed to 346 seeding devices (2 microfragments per device) for deployment.Individuals were evenly distributed among device treatments and orientation by genotype, and the devices were evenly seeded to the experimental sites.

Seeding device treatments and deployments
Reared spat and microfragments were deployed to Davies Reef (for site locations see Fig. 1a and Supplementary Table S2) in August 2021 and placed in 3 experimental treatments defined by the device type (Fig. 1d-f).Each device had nominally different levels of grazing exclusion and coral protection: (1) a featureless device acted as a positive control (open to grazing; Fig. 1d); (2) a caged featureless device acted as a negative control (complete exclusion of grazing by large fishes; Fig. 1f); and (3) a device with engineered fish-exclusion features (i.e., protrusions) acted as the experimental device (device of interest; Fig. 1e).The experimental device was designed at AIMS as a part of the Reef Restoration and Adaptation Program (RRAP) and represents the first design to support field deployment of juvenile corals grown on standard 20-mm (Ø) coral frag plugs.The devices were made of 95% alumina ceramic, which was selected for its chemical and physical stability (e.g., high specific gravity [3.8], hardness [9.0 Mohs]), and manufactured in the People's Republic of China (Shanghai Gongtao Ceramics CO., Ltd.PRC).The device design provided the option for controlled spatial positioning (i.e., in relation to the seafloor) and modular components where the device could be customized for specific experiments; in this case, we selected modules with and without fish exclusion features.
Within each experimental device, there were 6 positions for coral plugs spread among 2 orientations: top-and side-facing.In the side orientation, the fish exclusion device had 12 mm protrusions on two sides, with open access for grazing from the top and no access from the bottom (Fig. 1e and h).The top orientation had 12-mm protrusions on two sides, a 10-mm protrusion at the inner edge and no protection on the outer edge (Fig. 1e  and h).The minor differences in protection across plug orientations in the exclusion device were unavoidable due to assembly requirements but didn't appear to affect fish behavior (see results).The cage for the negative control was made from stainless steel metal fencing with a 25 × 25 mm grid (Fig. 1f).Devices were assembled and attached to the reef flat using a combination of metal strapping, bolts, nails, and cable ties, which were consistent among treatments (Fig. 1d-g).The central bolt used for device assembly provided an extra form of protection (i.e., physical barrier to large fish) to the inner edge of the coral plug on all devices (Fig. 1d-g).The devices were deployed onto 4 replicate sites at Davies Reef (Fig. 1a,b; Table S2) under GBRMPA permit G21/45,348.1 and were removed after 8 months.
The field deployment followed a hierarchical design (Fig. 1c) and included: sites (4, categorical, random, and fixed), coral life stage (2, categorical, fixed), device treatment (2 or 3, categorical, fixed), plug orientation (2, categorical, fixed) and replicates for each category.Sites (> 100 m apart) were located on the reef flat in 3m depth at high tide.Sites were selected based on their similarities in benthic community composition, their leeward location, and the presence of adult A. digitifera colonies.Within each site, devices (n = 76-100) were paired based on device treatment and coral life stage and deployed to 0.25 m 2 plots (n = 31-41; Fig. 1g,h).Plots were placed haphazardly within each site and spaced at least 2-m apart.Due to the limited number of plugs with sufficient coral spat at the time of deployment, the cage treatment was only tested with microfragments.Therefore, sites had plots with paired devices (featureless and exclusion, Fig. 1d-e and 1h) and plots with all 3 devices (Fig. 1d-g).Paired plots (n = 17-23 site −1 ) had spat and microfragment plugs (n = 2 corals life stage −1 , n = 1 orientation −1 , n = 4 device −1 ) while plots with 3 devices (n = 13-18) contained microfragments only (n = 1 orientation −1 , n = 2 device −1 ).In total, 346 seeding devices with 98 spat and 692 microfragment plugs were deployed.

Biological and environmental data collection
Devices were monitored in situ during 3 survey time points (2, 90, and 240 days) post deployment.Data collection included a suite of quantitative and qualitative assessments to determine the influence of biological (fish abundance and grazing, benthic composition) and environmental (reef hydrodynamics and sedimentation) drivers on coral growth and survival, each of which are detailed below.More details about the data collection for this experiment can be found in Supplementary Table S2.

Coral survival and grazing assessments
Coral plugs were imaged (Olympus TG6) by divers during each survey and images were assessed to determine the presence or absence of grazing marks (at time 2 d only) and surviving corals (all time points) at the level of coral plug.Corals in caged treatments were not imaged at 2 d due to the difficulty in removing and resecuring cages, and it was assumed that no grazing had occurred; corals in caged treatments were imaged at all survey timepoints thereafter.Grazing (presence or absence) was not assessed with imagery in subsequent timepoints due to the difficulty in distinguishing old from new bite marks on the plugs.Only devices that held live corals and remained fixed to the substrate for the entirety of the experiment (63 devices with spat and 275 devices with

Fish community surveys and surveillance
Fish communities were observed using in situ stationary point count surveys (78.5 m 2 for 5 min, 3 replicates per site) by divers at 3 timepoints (2, 90, and 240 d).Fish of the key grazing families (Labridae, Acanthuridae, Siganidae, Pomacentridae, Balistidae, and Chaetodontidae) were counted with total counts and relative abundance calculated for each family, timepoint and site.When possible, the same observer completed all surveys to reduce observer bias in the data.Observer 1 completed > 55% of the surveys.The time, depth, tide, sea state, visibility and reef complexity were also recorded during surveys.Sea state and reef complexity were scored based on descriptions from the AIMS Long Term Monitoring Program 52 .
Feeding and grazing frequency within a 0.25 m 2 plot around 2 haphazardly selected sets of seeding devices at each site were assessed using GoPro (HERO 9) video surveillance in the morning (8:30-11:30) and afternoon (14:00-17:00) of each survey timepoint (2, 90, and 240 d).GoPro cameras were mounted to lead dive weights using cable ties and left to record for 1-2 h in the absence of divers.The number of videos and hours of footage were standardized across sites and the fish grazing activity was recorded.The fish found to be feeding in the plots were identified to species and the following information was collected: number of feeding forays (attempts), number of bites per foray, device grazing (yes/no, per bite), plug grazing (yes/no, per bite), plot type (devices in pairs or triplicates depending on life-stage treatment), and device treatment (exclusion, featureless, or cage).Detailed information for video assessment criteria and data collection can be found in Supplementary Sect.1.3 and Table S3.

Benthic habitat and community assessments
Reef habitat assessments were undertaken at 4 spatial scales: plug, device, plot, and site level.Images of the coral seeding devices and plugs were taken at all timepoints to track the growth of benthic competitors across treatments and overtime.Within plots, 0.25 m 2 quadrats were used to estimate the cover of benthic community constituents (crustose coralline algae, turf algae, fleshy macroalgae [to genus], cyanobacteria, live hard coral [to genus], soft coral, other benthic invertebrates, invertebrate grazers, and 'bare' substrate [defined as recently grazed algal turfs present on hard rock or rubble]).Quadrats were placed centrally around the devices and imaged (Olympus TG6) during all 3 timepoints (2, 90 and 240 d; Table S2).Images were imported into the Reef Cloud online database (https:// reefc loud.ai/) and 25 randomly overlaid points per image were scored (human observer) to one of the benthic community categories.Image classifications (109-188 total images per site) were used for statistical analysis.
Site-level benthic data were obtained from in situ point-intercept surveys collected at the 2-d timepoint only (Table S2), from 3 replicate 30-m transects per site, with data recorded every 50 cm, allocated to the same benthic community constituent categories as above.During all 3 timepoints, a single 30-m video transect was recorded (Olympus TG6).Frame grabs (n = 35) were taken from video transects and scored (human observer) on Reef Cloud, with 10 randomly overlaid points per image.The in situ benthic data (at the site level) were compared to the image classifications (at the site level) and both data sets were investigated further using statistical analyses (see below).

Environmental data: wave energy and sediment
Clod cards (i.e., small blocks made of plaster of Paris) were used to compare relative water flow among sites by measuring dissolution over time and comparing Diffusion Factors (DF 53 , defined as the ratio between the weight of material dissolved in an experimental block to a control block maintained in a stable environment) 53,54 .Clod cards were glued to the lid of a plastic food container and secured to a lead dive weight with cable ties for deployment.Five replicate clod cards were pre-weighed, then deployed to each site for 48-72 h (Table S2).Upon retrieval, the food containers were gently placed over the clods and secured to the lids before returning them to the surface.Clod cards were dried and the weight loss (defined as the card value [CV] 53 , pre-minus postdeployment weight) was used to obtain the dissolution rate (g m −1 d −2 ).Weight loss was also used to calculate water velocity (cm s −1 ; V = [CV-5.8]/0.3) 53.Values for V were then used to obtain DF (DF = 0.06V + 1.3) 53 .To reduce variability in the data due to sea state, clod cards were deployed on the same day at all sites.
Deposited sediments were assessed using two methods, 'SedPods' and 'TurfPods' , that were created following techniques from Field et al. 2013 55 .Sediment collection pods (5 replicates per type) were deployed to sites (n = 4) at each survey timepoint (n = 3) and collected after 5-7 d (Table S2).From this, the total mass of deposited sediment was quantified for each pod type, which approximates sediment deposition on massive coral surfaces and algal turf-covered substrate, respectively.Pods were stored in a small volume of seawater at −20 °C and held until processing.Sediment processing followed standard water and wastewater methods from the American Public Health Association 2018 (see Supplementary Sec 1.4).Briefly, the individual samples were rinsed to eliminate salts (5-6 times with reverse osmosis water and a 3-h settlement period in between rinses), dried (24 h, 103-105 °C), and weighed to obtain the total mass per sample.Sediments were not separated by composition (organics or inorganics) or fractionated by particle size, due to the small mass obtained per sample.

Coral survival and predation from grazing fish
The coral survival and fish grazing data were analyzed using generalized linear mixed effect models in R statistical software 56 using the 'glmer' function in the 'lme4' package 57 .To investigate the change in survival through time and among treatments and life-history stages, we first modelled survival (binomial distribution with a Vol:.(1234567890 www.nature.com/scientificreports/logit link function) against the additive effects of survey time point, coral life stage, and the interaction between device treatment and plug position, with a random effect of site.To investigate whether survival was different among sites, we then modelled survival (at the final timepoint only) against the additive effect of site, device type and life stage as fixed effects, with other variables (plug position, and device number) as random effects.
Presence or absence of coral grazing after 2 d (binomial distribution with a logit link function) was modelled against coral life stage, site, and the interaction between device treatment and plug position as the fixed effects, with device number as the random effect.For each analysis, different combinations of predictors were tested, and the 'best' routine models were selected by comparing Akaike Information Criterion (AIC).Diagnostic plots of the residuals and Q-Q (DHARMa 58 ) were used to validate model assumptions and test for homogeneity of variance and linearity between the predictors and the response variables.All parameters were compared and tested for significance using the 'emmeans' package 59 and model outputs were plotted against the response for data visualization using the 'ggplot2' package 60 .

Fish abundance and grazing behavior across seeding sites
Redundancy analyses (RDAs, R software 56 , 'rda' function of the 'vegan' package 61 ) were used to explore the fish abundance and grazing data.From this, ordination plots were generated to identify the site or combinations of sites that explained the greatest differences in fish abundance and grazing intensity.Linear regression models (R software 56 , 'lm' function) were used to test the significance of fish families, genera, and species (abundance and grazing) against PC1 and PC2 of the RDAs.Additive combinations of species, genera and families were also tested.Likewise, site was regressed against PC1 and PC2 to test for its significance.Three zero-inflated generalized linear mixed effects models (R software 56 , 'glmmTMB' function and package 62 ) were used to further explore fish abundance and grazing by site and fish taxonomic group.First, fish abundances and bites (as the response variables) were modeled against site and fish categories (family-level identification) as predictors.Then, bites by parrotfish (Labridae, Scarini) were tested alongside site and species-level categories.A truncated Poisson distribution was used to accommodate count data that might exhibit overdispersion while still constraining the values to be non-negative.A zero-inflated component was included to address the excess zeros commonly observed in count data.

Ecological data: benthic composition, sedimentation, and wave energy
RDAs were conducted to investigate whether there was a clear separation in the benthic community composition among sites with higher and lower coral survival and grazing at the 2-d timepoint.The RDAs included abundance (percent cover of benthic community constituents per plot) against site, coral survival, or coral grazing as predictors, and visualized through ordination plots.A linear regression model was then used to investigate the relationship between PC1 and PC2, and the explanatory variables.Coral survival and grazing were also tested as the responses against PC1 and PC2 as additive predictors.Logistic regression models (glm) were used to test the abundance of significant benthic categories (groups or individual constituents) as predictors of coral survival and grazing.Models (lm and glm) were also built to investigate differences in wave energy and sedimentation among sites.Deposited sediments, diffusion rates, or diffusion factors were modeled against the predictors of site and pod type (for sediment data only).All models were run and validated as described above.

Results
Acropora digitifera survival and grazing significantly differed by device treatment, orientation, life stage and site.
Grazing pressure was also related to ecological attributes of the seeding site.Below we report the results of coral survival and grazing pressure, in turn, followed by the ecological variables that underpinned differences across sites.Please refer to Supplementary Tables S4 to S10 for the full results of statistical analyses.
Average survival was significantly higher in side-facing corals than top-facing ones, and this pattern was consistent across device treatments (featureless and exclusion feature), life-history stages, and through time    S5).Points represent the modelled estimated marginal means with 95% confidence intervals.Note that the x-axis in (a) is on a log scale.
(GLMM, p-value = 0.001, estimate = −0.65,SE = 0.20, z-ratio = −3.30;Table S4; Fig. 2a).This result was most pronounced for microfragments, with 1.5-times more survivors on side versus top oriented corals after 240 d.However, the best performing orientation and device combination for microfragments was the upward-facing plug in the caged treatment (Fig. 2a).Survival in these positions was 1.6-fold higher than upward-facing plugs in featureless device (lowest performing orientation and device combination) but was not significantly different from side-facing plugs in the exclusion device.After 240 d, there was no significant difference in spat or microfragment survival between sites (Fig. 2b).Site 1 had the highest number of surviving spat and microfragments at the end of the experiment, while Site 3 had the lowest (Fig. 2b); this result remained consistent across the device treatments.

Effects of fish grazing
Within 2 d of seeding, a significant number of corals on featureless devices were heavily grazed by fishes and were grazed up to 4-times more often than those in the exclusion treatment (GLMM, p-value < 0.001, estimate = −2.72,SE = 0.57, z-ratio = −4.78;Table S5; Fig. 2c).Top-orientated corals were targeted for grazing significantly more often (up to 18-times more) than side orientations (GLMM, p-value = 0.035, estimate = −0.83,SE = 0.39, z-ratio = −2.11;Table S5; Fig. 2c), and significantly more spat were grazed than microfragments (GLMM, p-value = 0.03, estimate = 0.77, SE = 0.36, z-ratio = 2.1; Table S5; Fig. 2c).Despite protection in the exclusion devices, up to 55% of spat and 25% of microfragments in the top orientation experienced grazing (Fig. 2c).However, the fish bites in exclusion devices were constrained to the outer edge of the top plug and therefore never caused 100% mortality.Overall, the device and orientation combination that experienced the least grazing was the side position of the exclusion device, while the most was the top position of the featureless device (GLMM, p-value = 0.01, estimate = −2.94,SE = 1.18, z-ratio = −2.5;Table S5; Fig. 2c).
Fish grazing was also site-specific.On average, corals had up to 5-times more grazing and predation attempts at Site 3 than Sites 1 and 2 (GLMM, p-value < 0.001, estimate = 3.48, SE = 0.71, z-ratio = 4.91; Table S5; Fig. 2c), regardless of device type, with up to 95% of spat and 75% of microfragments on featureless devices experiencing grazing there.
(c) Fish bites (min -1 ) from non-parrotfish species in plots, separated by site.Standard error and 95% confidence intervals are displayed for each site for the observed (c) and modeled (a, b) data, respectively.
Spat grazing and survival was also related to the abundance of Acropora corals with corymbose and digitate morphologies.For example, a 10% increase in Acropora corals was associated with a 10% decrease in the likelihood of grazing on spat and increased the chance of survival at Site 1 (Fig. S1).Site 1 had the highest overall cover of Acropora corals in plots, while Site 3 had the lowest.The influence Acropora coral cover on spat survival was most extreme when comparing Site 3 to Site 2 and 1, respectively (lm, Site 3 to 2: p-value = 0.01, estimate = −0.55,SE = 0.18, t-value = −3.03,and Site 3 to 1: p-value = 0.03, estimate = −0.28,SE = 0.11, t-value = -2.65;Table S9).The abundance of non-Acropora corals, octocorals, algal turfs and CCA contributed to site-related differences; however, not all variables correlated with coral grazing and/or survival, and the responses weren't consistent among sites (Table S9).
There was little to no change in the composition of benthic communities on plugs throughout the experiment (including caged devices).The dominant benthic category at the time of deployment (i.e., those that developed in ex-situ aquaria) remained dominant at the end of the experiment.
Environmental data revealed slight variation in water movement and sediment deposition at the site level.Overall, the range of water motion was low across sites at the 2-d timepoint (DF of 3.68 to 5.42 on a DF scale of 1 [low] to 20 [high]; Table S10; Fig. S3).The total mass of deposited sediments on Sed Pods explained more of the among-site variation than sediment accumulation on the Turf Pods.On average, the deposited sediments collected from Sed Pods at Site 1 were lower than sediments at other sites (Fig. 4d

Discussion
The experimental results indicated four ways to maximize survival of Acropora digitifera corals over an eightmonth seeding trial: (1) deploy devices with fish-exclusion features; (2) orient corals in the side rather than top positions; (3) use differential survival data to inform appropriate microfragment and spat seeding densities to achieve target numbers; and (4) use ecological variables, like fish abundance, fish diversity and the abundance of nutrient-dense food sources for fish, to help select seeding sites.These four findings can be used to guide coral seeding methods and manage restoration outcomes.Corals deployed on seeding devices without protection were the most vulnerable to grazing and suffered significantly higher mortality compared to those with protection.Complex device topographies, including crevices, microrefugia and those with high rugosity, have previously been shown to improve coral larval settlement and post-settlement survival 11,23,24,30 .Thus, it was not unexpected that the seeding device engineered with protective walls also enhanced coral survival after seeding.Although the caged device still resulted in higher survival of coral microfragments in our study the built-in protection provided by a fish-exclusion device offers a simple and effective alternative to reduce grazing pressure that is cheaper and far more scalable than cages.Furthermore, cages are ineffective against fish grazing after removal 33 .Nevertheless, there is room to improve the design of devices with features to offer more protection and further increase benefits over caged devices.
The orientation of corals in devices had a major influence on post-seeding survival.Coral larvae often preferentially settle on vertical surfaces to reduce the likelihood of disturbance-induced mortality during recruitment [63][64][65][66] .Our findings confirmed that corals (especially spat) deployed on top-facing plugs suffer higher rates of grazing and mortality than those in side-facing positions.Many species of parrotfish prefer to feed on horizontal and convex surfaces over vertical and concave surfaces 67 .Therefore, the vertical orientation of the side-facing plug may offer optimal refuge from some grazers.Our grazing results support the selection of vertical habitats by corals.However, there are also reasons beyond fish predation as to why coral larvae prefer settling in refugia.Coral spat are sensitive to high light, heavy sedimentation and excessive waterflow 68 , and verticallyorientated structures provide a buffer against such conditions.Fast growing turf, crustose, and macroalgae, that tend to out-compete corals in high-light and sediment-laden environments, can also be less abundant and display different morphologies (e.g., thin-versus thick-crusted CCA) in low-light environments 39,69 , relieving some competitive pressures.Regardless of the mechanisms, offering vertically oriented refugia for corals was a successful strategy for improving survival.
It was clear that coral life stage impacted survival after seeding, with significantly more A. digitifera microfragments surviving than spat.Spat survival was low (< 50%) on featureless devices past two days, and even with device protection.Low survival of spat is common in coral seeding experiments to date.For example, in French Polynesia, grazing led to mass mortality of A. striata spat after one-week of seeding 70 .Corals in cages had the highest survival but less than 40% survived beyond the initial week of deployment 70 .Similarly, in the Philippines, survival of A. tenuis spat was < 25% three-months post seeding and spat in partial cages outperformed those in full cages in this experiment 28 .By contrast, on the GBR, 38-65% of A. cytherea spat survived on reefs for one month, with caging increasing survivorship by 22% 27 .Finally, partial protection of A. tenuis spat in microrefugia led to 22-39% yield (e.g., device-level survival) after one year on the GBR 11,12 .Although these examples demonstrate that protection from fishes can increase survival during early ontogeny, spat survival typically remains well below 50%.We suggest that: (i) more spat per device need to be deployed to meet required survival outcomes, (ii) other device types should be trialed to improve the design of protective features, and (iii) further ecological drivers should be studied to inform optimal conditions for spat success after seeding.
While microfragmentation has almost exclusively been performed on corals with plating, massive and submassive growth morphologies 18,19 , our results indicate that the technique is also feasible when applied to branching taxa such as Acropora.Survival of A. digitifera microfragments was exceptionally high, greater than 75% on average and up to 97% in cages.These findings, alongside the result of previous experiments 30 , confirm the benefits of physical barriers to enhance the survival of coral microfragments.In Palau, small fragments of Porites lobata had higher survival when fully protected (> 90%, four-sided crevices on tiles) compared to the result of partial crevices (70%) and exposed controls (28%) after 29 days 30 .In the same study, Pocillopora damicornis fragments also initially benefited from protection; however, all had died after eight days.These outcomes indicate that tolerance to microfragmentation may be species specific and this will have flow-on effects to survival.Similarly, other factors including fragment health might dictate the likelihood of predation post deployment, with recently dead or dying tissues being more attractive to fishes 71 .The A. digitifera tested in our study may be more tolerant to microfragmentation and less palatable to fishes, making it a target candidate for seeding over other taxa such as Pocillopora.To ensure positive outcomes after seeding, adequate time should be provided for corals to heal after the microfragmentation process, and the palatability of a range of coral species should be examined.
We found that coral spat were also targeted for grazing more often than microfragments.Parrotfish, mainly Scarus globiceps, were identified as the most common and destructive grazers of seeded corals, especially spat.However, most scraping parrotfish are nominally herbivorous and therefore we hypothesize that the grazinginduced mortality of coral spat in our study is likely accidental and indirect.The morphological description of the jaw of a scraping parrotfish like S. globiceps, confirms their ability to graze early successional bacterial-algal biofilms growing on reef substrata 67,72 .The artificial plugs used to grow corals in our study were conditioned with similar biofilms (e.g., thin-crusted CCA, microalgae and mixed bacteria) and these are known to be palatable to many herbivorous parrotfishes 73,74 .Thus, it is likely parrotfish were targeting the benthic community growing on the plug rather than the coral itself.Furthermore, spat plugs were held in aquaria longer than microfragment plugs prior to deployment; this likely contributed to significant differences in grazing among the coral life stage treatment.The use of less palatable benthic organisms for plug conditioning and the incorporation of an antifoulant (i.e., chemical deterrent) on plugs or devices are potential direct and indirect solutions to deter parrotfish grazing.However, we suggest these methods must be balanced against the requirement for benthic communities that: (i) are inductive for the larvae during settlement, (ii) reduce competition against spat and microfragments during recruitment, and (iii) promote coral survival to maturity.
Lastly, the influence of grazing on survival is strongly site-specific, and this information may be useful to guide the selection of sites for successful seeding.Foremost, our results support growing evidence that herbivorous parrotfish may preferentially target nutrient-rich food sources (e.g., algal mats or turfs dominated by cyanobacteria), when available 75,76 .However, on reefs with high coral cover and low abundance of cyanobacteria, opportunistic grazing on less preferred food sources such as deployed corals may occur.For example, Site 2 exhibited the least grazing on coral plugs but the highest number of parrotfish bites in experimental plots, and concurrently, a notably higher (12 times) abundance of mat-forming cyanobacteria.Parrotfish consumption of nutrient-dense foods, such as endolithic algae and cyanobacteria, has been documented in the Caribbean 75 and the northern GBR 76 .Field observations performed by Cissell et al.(2019) found that greater than 15% of the diet of parrotfishes S. iseri and S. coeruleus consisted of mat-forming cyanobacteria 75 .Similarly, stable isotope analysis of 22 species of GBR-dwelling parrotfish provided strong evidence to support the consumption and preference of microscopic cyanobacteria over other algal substrates during feeding 76 .From this, we propose a new hypothesis that deployment adjacent to nutritious foods like cyanobacteria can offer natural forms of refuge to corals from grazing after seeding.
En masse, deploying to sites with high-nutrient food sources for fish may limit both direct and indirect grazing pressure on corals, increasing the likelihood of successful restoration.However, the composition of modern reefs is expected to shift as the frequency of mild to severe disturbances increases over time.For example, sudden spikes in the abundance of mat-forming cyanobacteria are common characteristics of recently disturbed reefs 77,78 .We suspect slight increases in cyanobacteria (< 20% in 0.25 m 2 ) can aid coral survival after seeding; however, too much cyanobacteria can negatively impact coral dominance on disturbed reefs.For A. digitifera spat, we found that a minimum cover of closely related Scleractinian corals (10%) and bare rock (20%) was correlated with a reduction in grazing by 10 to 50%, respectively.Indeed, other benthic constituents, like sponges and epilithic algal communities, have been correlated with high survival of seeded corals on the GBR 12 .Therefore, a diverse group of grazing fishes at a minimum biomass may be required to mediate competition between corals and fastgrowing benthic organisms after seeding.A relatively low biomass of herbivores (177 kg ha [17.7 g m 2 ]) has been shown to sustain community equilibrium and reduce the risk of regime shifts occurring in the Indian Ocean 79 .Defining a biomass threshold for parrotfish (e.g., where the net benefit towards coral competition outweighs the negative effects of accidental grazing), represents one tool that can be used to guide or tailor deployments of corals to specific sites.The direct manipulation of herbivore abundance (i.e., enhancing fish recruitment) could also be considered at sites with significant regime shifts.Taken together, these results highlight the importance of a holistic approach that considers multiple ecological parameters to inform the selection of candidate sites for reef restoration with coral seeding devices.

Conclusion
The addition of fish-exclusion features in coral seeding devices can improve survival and lead to substantial reductions in the handling time required for coral seeding, supporting the goal of upscaling restoration initiatives.Our eight-month seeding trial with Acropora digitifera revealed significantly higher survival when spat and microfragments were deployed in vertical positions with protection from grazing fishes.Importantly, future coral seeding experiments need to consider biological and environmental characteristics of the receiving environment more holistically and test across ecologically important reef-building coral taxa.Here we've identified that environmental drivers including a diverse assemblage of fishes (Labrids, Pomacentrids, Acanthurids, Chaetodontids) and low to moderate cover (< 40%) of coral, recently grazed reef rock, and nutrient-rich foods for parrotfish, could be used as visual aids (i.e., during pre-deployment assessments) to optimize the selection of sites for seeding in the future.While the role that herbivorous fishes play in driving coral-seeding success requires further investigation, our results suggest that restoration research can be used both to guide the development of coral seeding techniques and to advance our fundamental understand of reef ecology.

Figure 1 .
Figure 1.Site selection and design plan for the coral-seeding experiment.A total of 346 seeding devices were deployed to Davies Reef (a; Google Earth Image © 2023 Maxar Technologies) and fixed to the reef substrate at 4 sites (b; Google Earth Image © 2023 Maxar Technologies; see supplementary Tabe S2 for coordinates).The experimental design (c) included 2 coral life stages, 3 device treatments, and 2 plug orientations, with replication among categories.The device treatments (d-f) included featureless control devices (+ control; d), devices with fish-exclusion features (experimental treatment; e) and caged control devices (-control; f).Devices (n = 76-100) were paired based on device treatment and coral life stage and deployed to 0.25 m 2 plots (n = 31-41; g-h) at each site (n = 4).

Figure 2 .
Figure 2. Average coral survival and grazing on seeded Acropora digitifera.(a) Coral survival through time by device treatment, coral orientation, and coral life stage (n obs = 2456, n groups = 4; see Supplementary TableS4).(b) Coral survival at 240 d separated by site, device type, and life stage (n obs = 614, n groups = 2).(c) Grazing by fish on seeded corals at 2 d allocated by site, device treatment, orientation, and life stage (n obs = 380, n groups = 149; TableS5).Points represent the modelled estimated marginal means with 95% confidence intervals.Note that the x-axis in (a) is on a log scale.
Figure 2. Average coral survival and grazing on seeded Acropora digitifera.(a) Coral survival through time by device treatment, coral orientation, and coral life stage (n obs = 2456, n groups = 4; see Supplementary TableS4).(b) Coral survival at 240 d separated by site, device type, and life stage (n obs = 614, n groups = 2).(c) Grazing by fish on seeded corals at 2 d allocated by site, device treatment, orientation, and life stage (n obs = 380, n groups = 149; TableS5).Points represent the modelled estimated marginal means with 95% confidence intervals.Note that the x-axis in (a) is on a log scale.
Figure 2. Average coral survival and grazing on seeded Acropora digitifera.(a) Coral survival through time by device treatment, coral orientation, and coral life stage (n obs = 2456, n groups = 4; see Supplementary TableS4).(b) Coral survival at 240 d separated by site, device type, and life stage (n obs = 614, n groups = 2).(c) Grazing by fish on seeded corals at 2 d allocated by site, device treatment, orientation, and life stage (n obs = 380, n groups = 149; TableS5).Points represent the modelled estimated marginal means with 95% confidence intervals.Note that the x-axis in (a) is on a log scale. https://doi.org/10.1038/s41598-024-64294-z

Figure 3 .
Figure 3. Site-level abundance and bite rates for common reef fishes.(a) Counts of fish from in situ stationary point count surveys and grouped by fish family (n obs = 354, n groups = 11; see Supplementary TableS6).(b) Feeding (bites min -1 ) by parrotfish, Labridae (Scarini), in 0.25 m 2 plots within sites (n obs = 8040, n groups = 134; TableS7).(c) Fish bites (min -1 ) from non-parrotfish species in plots, separated by site.Standard error and 95% confidence intervals are displayed for each site for the observed (c) and modeled (a, b) data, respectively.

Figure 4 .
Figure 4. Ecological data collected across sites, compared to coral grazing and survival.Logistic relationships between the cover of bare substrate (a) or cyanobacteria (b) and the grazing of coral spat and microfragments by fishes at the 2-d timepoint.(c) Percent cover of mat-forming cyanobacteria recorded in device plots (0.25 m 2 , n = 31-41) in sites (n = 4) at the 2-d timepoint.(d) Sediment deposition (mg m -2 d -1 ) collected from Sed Pods (n = 15) deployed to sites (n = 4).